This project focuses on two serine threoninc protein kinases, GCK and GCKR and a tyrosine kinase PYK2. GCK and GCKR belong to a subfamily of protein kinases, which also includes GLK, HPK1, and NIK. They are characterized by an N-terminal kinase domain related to the yeast STE20 protein kinase, a large C-terminal regulatory domain, and the ability to activate the stress-activated protein kinase (SAPK, also referred to as Jun kinase or JNK) pathway. This pathway is activated by many cellular stresses, the inflammatory cytokine tumor necrosis factor (TNF) and CD40 ligand (CD40L). CD40L is present on activated T lymphocytes and interacts with its counter-receptor CD40, which is expressed by many on the important cells in the immune system, including dendritic cells, monocytes, and B-lymphocytes. Previously we showed that GCK and GCKR are major intermediaries in CD40 and TNF-mediated SAPK activation, that signaling through these receptors leads to the formation of a trimolecular complex between GCKR, TRAF2, and TANK. In conjunction with Z. Ronai we have shown that an increase in GCK and TRAF2 expression occurs during melanoma progression, which impacts the sensitivity of melanoma cells to UV-induced cell death. A screen for molecules that bind GCKR has resulted in the identification of a panel of strong interacting proteins that includes a number of adaptor proteins such as Crk and CrkL as well as several proteins that participate in the Ras-signaling pathway. To better understand the role of GCK and GCKR in vivo, the murine GCK and GCKR gene have been isolated. Both genes are complicated spanning nearly 100 kilobases with more than 30 exons. This information is being used to create gene targeting constructs. The tyrosine kinase PYK2 links signals generated by g-protein coupled receptors to downstream signaling pathways. One type of G-protein, G13alpha, potently activates PYK2 kinase activity, which leads to the activation of a small GTPase called RhoA, which in turn stimulates morphological changes in cells and triggers a conserved signaling pathway important for cell growth. The activation of g-proteins in the cells leads to a transient association between PYK2 and G13alpha. Whether G13alpha directly activates PYK2 or uses an intermediatory has not yet been resolved. Finally, in conjunction with J. Tuscano we have shown a potential role for antibodies against the B lymphocyte cell surface molecule CD22 in inducing apoptosis of human B cell lymphomas.